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NELFE / Negative elongation factor E · IHC design guide
Design Immunohistochemistry for NELFE
Plan paraffin-section NELFE IHC around mainly nuclear staining in most tissues (HPA tissue IHC). Use adrenal glandular cells or marrow hematopoietic cells as positive controls (HPA tissue IHC), and start the catalog antibody at 2–5 μg/ml (datasheet A07050-1).
Evidence assembled Oct 2026 · For research use; verify linked source records and product datasheet before use
NELFE Immunohistochemistry Experimental Design Guide
Expected localisation, validated protocols, controls and antibodies — the at-a-glance facts below, then the full design guide.
Must know before staining
| Expected localisation | Mainly nuclear in tissue (HPA tissue IHC) | |
| Staining pattern | Mainly nuclear staining in most tissues (HPA tissue IHC) | |
| Antigen retrieval | EDTA pH 8.0 HIER, heat-mediated (datasheet A07050-1) | |
| Positive control | Adrenal gland+4 more · see all | |
| Negative control | Heart muscle+4 more · see all |
Important caveats
Reasons your staining may differ from the expected pattern.
| Fixation | Keep fixation consistent across compared paraffin sections. (selected-SKU IHC image A07050-1) | |
| Caveat | Cardiomyocytes and skeletal myocytes: not detected (HPA tissue IHC) | |
| Regulation | P-TEFb promotes release from chromatin (UniProt) | |
| Isoform / epitope | 3 isoforms; map the epitope before isoform scoring (UniProt) |
Section 1
Recommended NELFE IHC & IF Protocols
The catalog antibody uses EDTA pH 8.0 heat retrieval (datasheet A07050-1). Four published NELFE IHC protocols provide additional starting points (PMC6831195; PMC7851912; PMC8290256; PMC5539779).
| Sample | Paraffin-embedded human lung squamous cell carcinoma tissue; fixative not specified (datasheet A07050-1) |
| Fixation | Image fixative and duration unreported (datasheet A07050-1); verify before use. |
| Sectioning | 4–5 µm sections on charged slides (standard) |
| Deparaffinisation | Xylene, graded ethanol series to water (standard) |
| Antigen retrieval | Heat retrieval: EDTA pH 8.0 (datasheet A07050-1); 20 min, 95–100 °C (standard) |
| Peroxidase block | 3% H2O2, 10 min, room temperature (standard) |
| Blocking | 10% goat serum (datasheet A07050-1) |
| Primary antibody | Rabbit anti-NELFE, 2-5 μg/ml (datasheet A07050-1) |
| Primary incubation | Overnight at 4 °C (datasheet A07050-1) |
| Detection | HRP-conjugated secondary, DAB chromogen (datasheet A07050-1) |
| Counterstain | Hematoxylin, blue, dehydrate and mount (standard) |
| Expected result | NELFE-positive staining in glandular cells of adrenal gland (HPA tissue IHC: High). HPA tissue profile: Mainly nuclear expression in most tissues. No signal in the no-primary control. |
💡Decision noteStart with EDTA pH 8.0 heat retrieval for the catalog antibody (datasheet A07050-1). Assess nuclear staining with matched controls (UniProt P18615 localisation).
Section 2
What Is the Expected NELFE Staining Pattern?
NELFE should appear mainly in nuclei across many tissues, with strong staining in cell populations such as adrenal glandular cells, bone marrow hematopoietic cells and cerebral cortex neurons (HPA: mainly nuclear; High in these cells). Chromatin association supports this location (UniProt P18615: nucleus and chromosome). It has no transmembrane segment (UniProt P18615: topology). HPA rates the tissue IHC evidence Enhanced, while reporting medium consistency with RNA expression (HPA: reliability).
| Distinct nuclear staining in adrenal glandular cells or bone marrow hematopoietic cells (HPA: High in both). | This fits the reported tissue pattern and nuclear location (HPA: tissue IHC; UniProt P18615: nucleus). Judge staining in the identified cells, rather than treating every cell on the section as a positive control (general IHC practice). |
| Predominantly cytoplasmic, membranous or extracellular staining, with little nuclear signal. | This conflicts with NELFE’s reported nuclear pattern and lack of a transmembrane segment (HPA: mainly nuclear; UniProt P18615: topology). Check the negative control and antibody specificity before interpreting the signal as NELFE (general IHC practice). |
| Strong signal in cardiomyocytes, skeletal myocytes or smooth muscle cells (HPA: Not detected in each). | These cell populations are poor positive controls in the HPA tissue images (HPA: Not detected). Unexpected staining warrants checks for cross-reactivity or endogenous detection activity; it does not alone prove either cause (general IHC practice). |
| Haze spreads across nuclei, cytoplasm and tissue spaces without clear cell boundaries. | A diffuse pattern cannot establish the mainly nuclear distribution reported for NELFE (HPA: tissue IHC). Review background in the no-primary control, blocking, detection and counterstain before scoring cells (general IHC practice). |
| No nuclear signal in adrenal glandular cells despite interpretable tissue morphology (HPA: High). | The expected positive population has failed to stain (HPA: High in adrenal glandular cells). First assess the run’s positive control, reagent performance and retrieval conditions; one blank section does not establish biological absence (general IHC practice). |
💡Expected NELFE appearanceA convincing positive shows clearly localized nuclear chromogen, potentially strong in adrenal glandular cells or bone marrow hematopoietic cells (HPA: mainly nuclear; High in both); isolated cytoplasmic or membranous color without nuclear staining is suspicious (HPA: mainly nuclear; UniProt P18615: topology).
| Nuclear distribution and chromatin association (HPA: mainly nuclear; UniProt P18615: chromatin). | Score signal against nuclear morphology and counterstain (general IHC practice). NELFE can leave chromatin after P-TEFb phosphorylation, so a chromatin association does not require a uniform nuclear texture (UniProt P18615: chromatin release). |
| Choice of tissue control (HPA: tissue IHC). | Adrenal glandular, bone marrow hematopoietic, breast glandular, bronchial respiratory epithelial and cerebral cortex neuronal cells are reported High; cardiomyocytes and skeletal myocytes are Not detected (HPA: tissue IHC). Match the control to its cell population. |
| Expression evidence and assay limits (UniProt P18615: tissue specificity; HPA: tissue IHC). | UniProt reports expression in heart and skeletal muscle, while HPA reports no IHC detection in cardiomyocytes or skeletal myocytes (UniProt P18615: tissue specificity; HPA: Not detected). Do not convert those cell-level IHC results into claims that the organs lack NELFE. |
| Antibody validation and isoforms (HPA: antibody validation; UniProt P18615: isoforms 1–3). | HPA007187 has Enhanced IHC validation (HPA: antibodies). Three isoforms are listed, but the supplied sources do not map antibody epitopes or establish isoform-specific IHC staining (UniProt P18615: isoforms; HPA: antibodies). |
| IF/ICC Q&A: What localisation should an IF image show? | Mainly nucleoplasm and nuclear bodies (HPA: ICC-IF, both enhanced). HPA lists Enhanced ICC validation for HPA007594 and HPA046502; those ICC results do not establish an IHC-P protocol for either antibody (HPA: antibodies). |
| Situation | Likely cause | Next action |
|---|---|---|
| No staining in a positive-control adrenal glandular population (HPA: High). | The run may have failed, or the chosen assay conditions may be unsuitable (general IHC practice); target-specific fixation sensitivity is unreported in the supplied sources. | Check control tissue, primary and detection reagents, and the run’s documented IHC-P retrieval conditions before calling the sample negative (general IHC practice). |
| Nuclei stain weakly in esophageal squamous epithelium or prostate glandular cells (HPA: Low). | Weak staining may reflect the reported lower tissue signal (HPA: Low in these cells), although assay performance still needs a control (general IHC practice). | Compare the same run with a reported High population and score the relevant cell type separately (HPA: tissue IHC; general IHC practice). |
| Color is predominantly cytoplasmic or membranous. | The compartment conflicts with reported nuclear localisation and topology (HPA: mainly nuclear; UniProt P18615: no transmembrane segment). | Inspect the no-primary control and repeat specificity checks with an IHC-validated antibody before assigning the color to NELFE (general IHC practice; HPA: HPA007187 Enhanced IHC). |
| Cardiomyocytes or skeletal myocytes stain strongly (HPA: Not detected). | Cross-reactivity or endogenous detection activity is possible, but the image alone cannot identify which (general IHC practice). | Compare negative controls and the expected nuclear pattern in a High cell population; investigate the detection system if color persists without primary antibody (HPA: tissue IHC; general IHC practice). |
| Diffuse color obscures nuclear boundaries. | Nonspecific background or excessive detection signal may obscure the reported nuclear pattern (general IHC practice; HPA: mainly nuclear). | Review no-primary background, blocking, primary dilution, chromogen development and counterstain using the documented IHC workflow (general IHC practice). |
| A proposed IF result is used to justify an IHC-P call. | HPA reports nucleoplasm and nuclear bodies by ICC-IF, while the listed Enhanced ICC antibodies differ from the Enhanced IHC antibody (HPA: subcellular; HPA: antibodies). | Interpret the chromogenic section against tissue IHC and its IHC-validated antibody; use IF localisation as supporting context only (HPA: tissue IHC; HPA: antibodies; general IHC practice). |
Experiment evidence
IHC & IF images showing NELFE staining
A07050-1 · IHCIHC analysis of NELFE using anti-NELFE antibody (A07050-1). NELFE was detected in a paraffin-embedded section of human lung squamous cell ca
A07050-1 · IHCIHC analysis of NELFE using anti-NELFE antibody (A07050-1). NELFE was detected in a paraffin-embedded section of human ovarian cancer tissue
A07050-1 · IFIF analysis of NELFE using anti-NELFE antibody (A07050-1) and anti-Beta Tubulin antibody (M01857-3). NELFE was detected in immunocytochemica
HPA · Cerebral cortexHuman Protein Atlas IHC, Cerebral cortex. Source: https://www.proteinatlas.org/ENSG00000204356-NELFE/tissue/cerebral+cortex#imid_2358516
HPA · ColonHuman Protein Atlas IHC, Colon. Source: https://www.proteinatlas.org/ENSG00000204356-NELFE/tissue/colon#imid_2358442
HPA · LiverHuman Protein Atlas IHC, Liver. Source: https://www.proteinatlas.org/ENSG00000204356-NELFE/tissue/liver#imid_2358443
HPA · KidneyHuman Protein Atlas IHC, Kidney. Source: https://www.proteinatlas.org/ENSG00000204356-NELFE/tissue/kidney#imid_2358444
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA046502; green = NELFE, blue = nucleus, red = microtubules.
HPA · ICC-IF A-431Human Protein Atlas immunofluorescence, A-431, antibody HPA046502; green = NELFE, blue = nucleus, red = microtubules.
← swipe / scroll for more images →
Real IHC / IF data behind the staining-pattern expectations in this section.
Sample controls for NELFE IHC & IF
🧪Run breast first and score its glandular cells for nuclear NELFE staining (HPA: High in breast glandular cells; UniProt P18615: nucleus). Use skeletal muscle myocytes as the negative tissue (HPA: Not detected in skeletal muscle myocytes); on the breast slide, any cells without specific nuclear signal should show counterstain alone, but their target-negative status requires separate validation (HPA: High is assigned to glandular cells only).
Positive control tissue: Adrenal gland (Glandular cells, HPA High)
Negative control tissue: Heart muscle (HPA Not detected)
ICC-IF cell lines (HPA subcellular resource): HPA ICC-IF images show NELFE in A-431, U-251MG, U2OS, Rh30, SH-SY5Y, NIH 3T3, with annotated localisation: Nucleoplasm (enhanced), Nuclear bodies (enhanced) (HPA subcellular).
Technical controls: Include a no-primary, secondary-only control and a concentration-matched rabbit IgG isotype control (selected-SKU caption: rabbit primary and goat anti-rabbit secondary). A NELFE knockout specimen provides a biological negative control (standard IHC practice).
Quench endogenous peroxidase and check for residual DAB background on the breast section (standard chromogenic IHC practice).
⚠️Feasibility: A target-specific fixation window and fixation effect are unreported, and the selected-SKU paraffin-section caption does not state the fixative (selected-SKU caption: fixative unreported). The caption uses heat retrieval in EDTA at pH 8.0 for a lung carcinoma section, which offers a starting condition but does not establish retrieval dependence in breast (selected-SKU caption: EDTA heat retrieval in paraffin-embedded lung carcinoma). IF/ICC may make nuclear localization easier to assess because HPA reports nucleoplasmic and nuclear-body signal, while frozen-section performance is unreported; breast DAB scoring should account for endogenous peroxidase background (HPA: nucleoplasm and nuclear bodies; standard IHC practice).
HPA tissue IHC evidence for NELFE
Comprehensive Human Protein Atlas IHC scoring per tissue (reliability: Enhanced — Medium consistency between antibody staining and RNA expression data.). Rows are taken directly from the HPA tissue chart — click any row's HPA link to view the source.
Positive expression · recommended positive controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Adrenal gland | Glandular cells | High | Protein (IHC) | HPA → |
| Bone marrow | Hematopoietic cells | High | Protein (IHC) | HPA → |
| Breast | Glandular cells | High | Protein (IHC) | HPA → |
| Bronchus | Respiratory epithelial cells | High | Protein (IHC) | HPA → |
| Cerebral cortex | Neuronal cells | High | Protein (IHC) | HPA → |
Undetected expression · recommended negative controls
| Tissue | Cell type | Level | Evidence | Source |
|---|---|---|---|---|
| Heart muscle | Cardiomyocytes | Not detected | Protein (IHC) | HPA → |
| Parathyroid gland | Glandular cells | Not detected | Protein (IHC) | HPA → |
| Skeletal muscle | Myocytes | Not detected | Protein (IHC) | HPA → |
| Smooth muscle | Smooth muscle cells | Not detected | Protein (IHC) | HPA → |
| Soft tissue | Peripheral nerve | Not detected | Protein (IHC) | HPA → |
Section 3
Advanced NELFE IHC Tips
Troubleshoot NELFE staining in paraffin sections by checking retrieval, nuclear localisation, controls and scoring before interpreting differences between specimens.
Which retrieval condition should I try first for weak NELFE staining?
Start with heat-mediated EDTA retrieval at pH 8.0 for paraffin sections (datasheet A07050-1). The selected tissue image used this condition, followed by 2 μg/ml primary antibody overnight at 4°C, so check that the full workflow was followed before changing retrieval (datasheet A07050-1). If nuclear staining remains weak, compare a longer or shorter heating step on adjacent sections while keeping the antibody concentration and detection conditions fixed. Assess whether signal improves within nuclei rather than at section edges or damaged areas, since NELFE is mainly nuclear in tissue IHC (HPA: tissue IHC profile).
Could fixation explain inconsistent NELFE staining between paraffin blocks?
Target-specific fixation sensitivity is unknown: the selected paraffin-section caption does not state a fixative, and its staining cannot establish how NELFE responds to fixation (datasheet A07050-1). Record each block’s fixative and fixation duration, then compare adjacent sections processed with the same EDTA pH 8.0 retrieval and 2 μg/ml primary antibody conditions (datasheet A07050-1). Examine nuclear detail and tissue preservation alongside staining, because poor morphology makes a weak signal difficult to interpret. If blocks differ in processing history, use a matched reference section in each run before attributing a staining difference to NELFE abundance.
How should I troubleshoot cytoplasmic or uneven nuclear NELFE staining?
NELFE staining should be assessed primarily in nuclei: tissue IHC is mainly nuclear, while cellular imaging places the protein in nucleoplasm and nuclear bodies (HPA: tissue IHC profile; HPA: subcellular localisation). Chromatin association is also reported, and P-TEFb phosphorylation promotes release from chromatin (UniProt P18615: PubMed:14701750). Compare nuclear signal with the counterstain across intact cells, then inspect cytoplasmic colour in a no-primary control before calling it specific. Distinguish uneven signal within preserved nuclei from staining confined to torn edges, folds or necrotic regions, and repeat the run if those artefacts dominate the section.
Could isoforms or epitope masking account for discordant NELFE staining?
NELFE has 3 annotated isoforms, an RNA recognition motif at residues 262–332, and multiple modified residues, including phosphoserines (UniProt P18615). The supplied evidence does not map the catalog antibody’s epitope or establish which isoforms it recognises, so staining differences cannot be assigned to a particular variant or modification. First compare sections using the documented EDTA pH 8.0 retrieval and 2 μg/ml primary concentration (datasheet A07050-1). If results remain discordant, obtain epitope and isoform coverage information for the antibody and confirm the pattern with an independently characterised reagent before making an isoform-specific claim.
How can I assess NELFE by multiplex immunofluorescence alongside tissue IHC?
Treat IF as a separate validation exercise: the supplied staining workflow demonstrates paraffin-section chromogenic IHC for A07050-1, while HPA reports NELFE in nucleoplasm and nuclear bodies (datasheet A07050-1; HPA: subcellular localisation). Pair nuclear NELFE signal with a marker for an expected cell population, such as bronchial respiratory epithelial cells, where tissue IHC reports high staining (HPA: bronchus). Choose a spectrally separated, preferably far-red fluorophore when tissue autofluorescence affects shorter wavelengths, and include single-colour controls. Because NELFE is nuclear and lacks a transmembrane segment, optimise permeabilisation for nuclear antibody access while preserving nuclear morphology (UniProt P18615: localisation and topology).
What should I check when DAB background obscures nuclear NELFE signal?
The selected paraffin-section workflow used 10% goat serum blocking, 2 μg/ml rabbit primary antibody overnight at 4°C, and peroxidase detection with DAB (datasheet A07050-1). Check that the peroxidase block and washes are effective, then inspect a no-primary section for colour caused by endogenous enzyme activity or detection reagents. Compare background in nuclei, cytoplasm and empty tissue spaces before adjusting primary concentration or incubation, changing one variable at a time. A diffuse deposit outside intact nuclei is less persuasive for NELFE than reproducible nuclear staining, given its mainly nuclear tissue profile (HPA: tissue IHC profile).
How should I score NELFE staining across specimens with different cell mixtures? ⚠ ANSWER MARKED FOR VERIFICATION
Score intact nuclei within a predefined cell population, since NELFE tissue staining is mainly nuclear and varies across reported cell types (HPA: tissue IHC profile). Record the percentage of positive nuclei and staining intensity, or calculate a nuclear H-score from 0–300; keep thresholds and imaging settings consistent across specimens. Normalise positive counts to the number of evaluable nuclei in that population, or report positive nuclei per mm² with the sampled area stated. Exclude folds, necrosis and section edges using the same criteria throughout, and report cell composition separately so a population shift is not mistaken for altered NELFE staining.
What evidence distinguishes genuine NELFE positivity from a staining artefact?
Look for reproducible nuclear signal in intact cells, consistent with mainly nuclear tissue IHC and nucleoplasmic or nuclear-body localisation (HPA: tissue IHC profile; HPA: subcellular localisation). Check the identity of stained cells: bronchial respiratory epithelial cells show high staining, whereas skeletal muscle myocytes were not detected in the cited tissue profile (HPA: bronchus; HPA: skeletal muscle). Distrust colour restricted to section edges, folds or necrosis, and use a no-primary control to assess endogenous peroxidase or detection background. Interpret an absent or unexpected pattern alongside morphology and a run-matched positive reference; the HPA tissue assessment reports only medium consistency between antibody staining and RNA expression (HPA: Enhanced reliability description).
Boster reagents
Best NELFE / Negative elongation factor E IHC Antibodies
A07050-1 has IHC data from human paraffin sections and IF/ICC data from A549 cells (catalog: IHC/IF image captions).
Real IHC data
Anti-NELFE Antibody ®
Cat # A07050-1
A07050-1 lists human reactivity and IHC, IF, and ICC applications (catalog: applications/reactivity). Its captions show paraffin-section IHC in human lung squamous cell carcinoma and ovarian cancer, plus IF/ICC in A549 cells (catalog: IHC/IF image captions).
Which to pick: Choose A07050-1 for human paraffin-section IHC; its tissue captions report EDTA retrieval at pH 8.0 and 2 μg/ml primary antibody (catalog: IHC image captions). The same SKU has IF/ICC data in A549 cells at 5 μg/ml; clonality is unreported. Reactivity is listed only for Human, so cross-species use is not established; the paraffin-section captions do not report a fixative (catalog: reactivity; catalog: IHC image captions).
Each figure is that product's own IHC / IF validation image from its datasheet.
References
- UniProt Consortium. UniProt entry P18615 (NELFE_HUMAN, Negative elongation factor E).
- Human Protein Atlas. NELFE tissue IHC expression (reliability: Enhanced).
- Human Protein Atlas. NELFE subcellular location (ICC-IF): Mainly localized to the nucleoplasm and nuclear bodies..
- Human Protein Atlas. NELFE antibody validation summary (3 antibodies).
- NELFE promoted pancreatic cancer metastasis and the epithelial‑to‑mesenchymal transition by decreasing the stabilization of NDRG2 mRNA. International journal of oncology 2019 — PMC6831195.
- Overexpression of NELFE contributes to gastric cancer progression via Wnt/β-catenin signaling-mediated activation of CSNK2B expression. Journal of experimental & clinical cancer research : CR 2021 — PMC7851912.
- The RNA-Binding Protein NELFE Promotes Gastric Cancer Growth and Metastasis Through E2F2. Frontiers in oncology 2021 — PMC8290256.
- Oncogenic Activation of the RNA Binding Protein NELFE and MYC Signaling in Hepatocellular Carcinoma. Cancer cell 2017 — PMC5539779.
- PubMed PMID:2612324 — UniProt-cited evidence.
- PubMed PMID:8373374 — UniProt-cited evidence.
- PubMed PMID:14656967 — UniProt-cited evidence.
Immunohistochemistry protocol sheet for NELFE (Negative elongation factor E). Expected localisation Mainly nuclear in tissue (HPA tissue IHC). Antigen retrieval: EDTA pH 8.0 HIER, heat-mediated (datasheet A07050-1). Validated IHC antibody: A07050-1 at 2-5 μg/ml (the product's own IHC validation image is shown). Controls (HPA): positive Adrenal gland (Glandular cells); negative Heart muscle. IHC-P workflow on paraffin sections; fixative unspecified. Protocol steps: 1. Sample: Paraffin-embedded human lung squamous cell carcinoma tissue; fixative not specified (datasheet A07050-1); 2. Fixation: Image fixative and duration unreported (datasheet A07050-1); verify before use.; 3. Sectioning: 4–5 µm sections on charged slides (standard); 4. Deparaffinisation: Xylene, graded ethanol series to water (standard); 5. Antigen retrieval: Heat retrieval: EDTA pH 8.0 (datasheet A07050-1); 20 min, 95–100 °C (standard); 6. Peroxidase block: 3% H2O2, 10 min , room temperature (standard); 7. Blocking: 10% goat serum (datasheet A07050-1); 8. Primary antibody: Rabbit anti-NELFE, 2-5 μg/ml (datasheet A07050-1); 9. Primary incubation: Overnight at 4 °C (datasheet A07050-1). Watch for: Fixation: Keep fixation consistent across compared paraffin sections. (selected-SKU IHC image A07050-1); Caveat: Cardiomyocytes and skeletal myocytes: not detected (HPA tissue IHC); Isoform / epitope: 3 isoforms; map the epitope before isoform scoring (UniProt); Regulation: P-TEFb promotes release from chromatin (UniProt). Evidence: UniProt P18615, HPA tissue IHC · Enhanced, HPA ICC-IF · Mainly localized to the nucleoplasm and nuclear bodies., Datasheet A07050-1, PMC6831195, PMC7851912. QR code links to https://www.bosterbio.com/bosterbio-gene-info-cards/NELFE/immunohistochemistry-ihc-guide.